ABR/DPOAE publications

Cisplatin-related ototoxicity is a critical side effect of chemotherapy and can lead to irreversible hearing loss. This study aimed to assess the potential effect of the DNA methyltransferase (DNMT) inhibitor RG108 on cisplatin-induced ototoxicity. Immunohistochemistry, apoptosis assay, and auditory brainstem response (ABR) were employed to determine the impacts of RG108 on cisplatin-induced injury in murine hair cells (HCs) and spiral ganglion neurons (SGNs). Rhodamine 123 and TMRM were utilized for mitochondrial membrane potential (MMP) assessment. Reactive oxygen species (ROS) amounts were evaluated by Cellrox green and Mitosox-red probes. Mitochondrial respiratory function evaluation was performed by determining oxygen consumption rates (OCRs). The results showed that RG108 can markedly reduce cisplatin induced damage in HCs and SGNs, and alleviate apoptotic rate by protecting mitochondrial function through preventing ROS accumulation. Furthermore, RG108 upregulated BCL-2 and downregulated APAF1, BAX, and BAD in HEI-OC1 cells, and triggered the PI3K/AKT pathway. Decreased expression of low-density lipoprotein receptor-related protein 1 (LRP1) and high methylation of the LRP1 promoter were observed after cisplatin treatment. RG108 treatment can increase LRP1 expression and decrease LRP1 promoter methylation. In conclusion, RG108 might represent a new potential agent for preventing hearing loss induced by cisplatin via activating the LRP1-PI3K/AKT pathway.

Cisplatin-induced ototoxicity can be partially attributed to excessive reactive oxygen species (ROS) production, and agmatine is well-known for the activation of the phosphoinositide-3-kinase (PI3K)/protein kinase B (AKT) pathway to inhibit ROS production. Whether agmatine could be used to alleviate cisplatin-induced ototoxicity is investigated. Cisplatin-exposed House Ear Institute-Organ of Corti 1 (HEI-OC1) cells and cochlear explants showed increased ROS production detected by 2',7'-dichlorodihydrofluorescein diacetate (DCFH-DA) staining and decreased cell viability detected by Cell Counting Kit-8 (CCK-8) or Myosin 7a staining, which could be reversed by the agmatine pretreatment. Cisplatin intraperitoneally injected C57BL/6 mice demonstrated damaged auditory function as indicated by distortion products otoacoustic emissions (DPOAEs) and auditory brainstem response (ABR) assays, and trans-tympanically administrated agmatine in the left ears could partly prevent the auditory function loss. Mechanistically, downregulated B-cell lymphoma 2 (Bcl-2) expression, upregulated Bcl2-associated x (Bax) expression, and diminished p-PI3K and p-AKT expression were detected in cisplatin-exposed HEI-OC1 cells and cochlear explants, which could be prevented by the pretreatment with agmatine. Our investigation demonstrates that agmatine pretreatment could alleviate cisplatin-induced ototoxicity with the activation of PI3K/AKT signaling pathway.

Background The transcription factor Sox2 plays important roles in the developmental processes of multiple organs and tissues. However, whether Sox2 can protect mature or terminally differentiated cells against injury is still unknown. Methods We investigated the roles of Sox2 in cochlear hair cells, which are terminally differentiated cells, using conditional transgenic mice and several hearing loss models. Results Sox2 overexpression dramatically mitigated the degree of cochlear hair cell loss when exposed to ototoxic drugs. Noise-induced apoptosis of cochlear hair cells and hearing loss were also significantly alleviated by Sox2 overexpression. Notably, noise-induced upregulation of pro-inflammatory factors such as TNF-α and IL6 was inhibited by Sox2 overexpression. Then we used lipopolysaccharide to clarify the effect of Sox2 on cochlear inflammation, and Sox2 overexpression significantly inhibited lipopolysaccharide-induced upregulation of pro-inflammatory factors and alleviated inflammation-related cochlear hair cell death. Conclusions These results demonstrate a novel protective role of Sox2 in mature and terminally differentiated cochlear hair cells by inhibiting inflammation.

The transcription factor c-Myb is vital for cell survival, proliferation, differentiation, and apoptosis. We have previously reported that c-Myb knockdown exacerbates neomycin-induced damage to cochlea cells. However, the function and regulation of c-Myb in the mammalian inner ear remains unclear. Here, we first found that the expression of c-Myb in cochlear HCs was downregulated after cisplatin damage in vivo. Next, to investigate the role of c-Myb in HCs treated with cisplatin, the recombinant virus AAV-ie-CAG-Myb-HA (AAV-c-Myb) that overexpresses c-Myb was constructed and transfected into HCs. The protein expression of c-Myb was effectively up-regulated in cultured cochlear HCs after the virus transfection, which increased cochlear HC viability, decreased HC apoptosis and reduced intracellular reactive oxygen species (ROS) levels after cisplatin injury in vitro. The overexpression of c-Myb in HCs after AAV-c-Myb transfection in vivo also promoted HC survival, improved the hearing function of mice and reduced HC apoptosis after cisplatin injury. Furthermore, c-Myb-HC conditional knockout mice (Prestin; c-Myb-cKO) in which c-Myb expression is downregulated only in cochlear OHCs were generated and the cisplatin-induced HCs loss, apoptosis and hearing deficit were all exacerbated in Prestin; c-Myb-cKO mice treated with cisplatin in vivo. Finally, mechanistic studies showed that upregulation of the PI3K/Akt signaling pathway by c-Myb contributed to the increased HC survival after cisplatin exposure in vitro. The findings from this work suggest that c-Myb might serve as a new target for the prevention of cisplatin-induced HC damage and hearing loss.

Aminoglycosides (AGs) are commonly used antibiotics that cause deafness through the irreversible loss of cochlear sensory hair cells (HCs). How AGs enter the cochlea and then target HCs remains unresolved. Here, we performed time-lapse multicellular imaging of cochlea in live adult hearing mice via a chemo-mechanical cochleostomy. The in vivo tracking revealed that systemically administered Texas Red–labeled gentamicin (GTTR) enters the cochlea via the stria vascularis and then HCs selectively. GTTR uptake into HCs was completely abolished in transmembrane channel-like protein 1 (TMC1) knockout mice, indicating mechanotransducer channel-dependent AG uptake. Blockage of megalin, the candidate AG transporter in the stria vascularis, by binding competitor cilastatin prevented GTTR accumulation in HCs. Furthermore, cilastatin treatment markedly reduced AG-induced HC degeneration and hearing loss in vivo. Together, our in vivo real-time tracking of megalin-dependent AG transport across the blood–labyrinth barrier identifies new therapeutic targets for preventing AG-induced ototoxicity.

Functional hair cell regeneration in the adult mammalian inner ear remains challenging. This study aimed to study the function of new hair cells induced by a DNA demethylating agent 5-azacytidine. Adult mice were deafened chemically, followed by injection of 5-azacytidine or vehicle into the inner ear. Functionality of regenerated hair cells was evaluated by expression of hair cell proteins, auditory brainstem response (ABR), and distortion-product otoacoustic emission (DPOAE) tests for 6 weeks. In the vehicle-treated group, no cells expressed the hair cell-specific protein myosin VIIa in the cochlea, whereas numerous myosin VIIa-expressing cells were found in the 5-azacytidine-treated cochlea, suggesting the regeneration of auditory hair cells. Moreover, regenerated hair cells were co-labeled with functional proteins espin and prestin. Expression of ribbon synapse proteins suggested synapse formation between new hair cells and neurons. In hearing tests, progressive improvements in ABR [5–30 dB sound pressure level (SPL)] and DPOAE (5–20 dB) thresholds were observed in 5-azacytidine-treated mice. In vehicle-treated mice, there were <5 dB threshold changes in hearing tests. This study demonstrated the ability of 5-azacytidine to promote the functional regeneration of auditory hair cells in a mature mouse model via DNA demethylation, which may provide insights into hearing regeneration using an epigenetic approach.

Cell cycle associated protein 1 (Caprin1) is an RNA-binding protein that can regulate the cellular post-transcriptional response to stress. It is a component of both stress granules and neuronal RNA granules and is implicated in neurodegenerative disease, synaptic plasticity and long-term memory formation. Our previous work suggested that Caprin1 also plays a role in the response of the cochlea to stress. Here, targeted inner ear-deletion of Caprin1 in mice leads to an early onset, progressive hearing loss. Auditory brainstem responses from Caprin1 -deficient mice show reduced thresholds, with a significant reduction in wave-I amplitudes compared to wildtype. Whilst hair cell structure and numbers were normal, the inner hair cell-spiral ganglion neuron (IHC-SGN) synapse revealed abnormally large post-synaptic GluA2 receptor puncta, a defect consistent with the observed wave-I reduction. Unlike wildtype mice, mild-noise-induced hearing threshold shifts in Caprin1 -deficient mice did not recover. Oxidative stress triggered TIA-1/HuR-positive stress granule formation in ex-vivo cochlear explants from Caprin1- deficient mice, showing that stress granules could still be induced. Taken together, these findings suggest that Caprin1 plays a key role in maintenance of auditory function, where it regulates the normal status of the IHC-SGN synapse.

Several studies identified hearing loss as a risk factor for aging-related processes, including neurodegenerative diseases, as dementia and age-related hearing loss (ARHL). Although the association between hearing impairment in midlife and ARHL has been widely documented by epidemiological and experimental studies, the molecular mechanisms underlying this association are not fully understood. In this study, we used an established animal model of ARHL (C57BL/6 mice) to evaluate if early noise-induced hearing loss (NIHL) could affect the onset or progression of age-related cochlear dysfunction. We found that hearing loss can exacerbate ARHL, damaging sensory-neural cochlear epithelium and causing synaptopathy. Moreover, we studied common pathological markers shared between hearing loss and ARHL, demonstrating that noise exposure can worsen/accelerate redox status imbalance [increase of reactive oxygen species (ROS) production, lipid peroxidation, and dysregulation of endogenous antioxidant response] and vascular dysfunction [increased expression of hypoxia-inducible factor-1alpha (HIF-1α) and vascular endothelial growth factor C (VEGFC)] in the cochlea. Unveiling the molecular mechanisms underlying the link between hearing loss and aging processes could be valuable to identify effective therapeutic strategies to limit the effect of environmental risk factors on age-related diseases.

Mammalian hearing involves the mechanoelectrical transduction (MET) of sound-induced fluid waves in the cochlea. Essential to this process are the specialised sensory cochlear cells, the inner (IHCs) and outer hair cells (OHCs). While genetic hearing loss is highly heterogeneous, understanding the requirement of each gene will lead to a better understanding of the molecular basis of hearing and also to therapeutic opportunities for deafness. The Neuroplastin ( Nptn ) gene, which encodes two protein isoforms Np55 and Np65, is required for hearing, and homozygous loss-of-function mutations that affect both isoforms lead to profound deafness in mice. Here we have utilised several distinct mouse models to elaborate upon the spatial, temporal, and functional requirement of Nptn for hearing. While we demonstrate that both Np55 and Np65 are present in cochlear cells, characterisation of a Np65-specific mouse knockout shows normal hearing thresholds indicating that Np65 is functionally redundant for hearing. In contrast, we find that Nptn -knockout mice have significantly reduced maximal MET currents and MET channel open probabilities in mature OHCs, with both OHCs and IHCs also failing to develop fully mature basolateral currents. Furthermore, comparing the hearing thresholds and IHC synapse structure of Nptn- knockout mice with those of mice that lack Nptn only in IHCs and OHCs shows that the majority of the auditory deficit is explained by hair cell dysfunction, with abnormal afferent synapses contributing only a small proportion of the hearing loss. Finally, we show that continued expression of Neuroplastin in OHCs of adult mice is required for membrane localisation of Plasma Membrane Ca 2+ ATPase 2 (PMCA2), which is essential for hearing function. Moreover, Nptn haploinsufficiency phenocopies Atp2b2 (encodes PMCA2) mutations, with heterozygous Nptn -knockout mice exhibiting hearing loss through genetic interaction with the Cdh23 ahl allele. Together, our findings provide further insight to the functional requirement of Neuroplastin for mammalian hearing.

Noise exposure causes noise-induced hearing loss (NIHL). NIHL exhibits loss of inner ear sensory hair cells and is often irreparable. Although oxidative stress is involved in hearing loss, the complex mechanisms involved in NIHL are unclear. Hypoxia-inducible factor 1α (HIF-1α) has been suggested to be essential for protecting sensory hair cells. Additionally, it has been shown that ROS is involved in modulating the stability of HIF-1α. To investigate the NIHL pathogenesis, we established a tert-butyl hydroperoxide (t-BHP)-induced oxidative stress damage model in hair-like HEI-OC1 cells and an NIHL model in C57BL/6 mice. Protein and mRNA expression were determined, and biochemical parameters including reactive oxygen species (ROS) accumulation, glucose uptake, adenosine triphosphat (ATP) production, and mitochondrial content were evaluated. In HEI-OC1 cells, t-BHP induced ROS accumulation and reduced mitochondrial content and oxygen consumption, but the ATP level was unaffected. Additionally, there was increased glucose uptake and lactate release along with elevated expression of HIF-1α, glucose transporter 1, and several glycolytic enzymes. Consistently, noise trauma induced oxidative stress and the expression of HIF-1α and glycolytic enzymes in mice. Thus, we concluded that ROS induced HIF-1α expression, which promoted glycolysis, suggesting a metabolic shift maintained the ATP level to attenuate hair cell damage in NIHL.

Congenital cytomegalovirus (cCMV) infection is the leading infectious cause of neurodevelopmental disorders. However, the neuropathogenesis remains largely elusive due to a lack of informative animal models. In this study, we developed a congenital murine CMV (cMCMV) infection mouse model with high survival rate and long survival period that allowed long-term follow-up study of neurodevelopmental disorders. This model involves in utero intracranial injection and mimics many reported clinical manifestations of cCMV infection in infants, including growth restriction, hearing loss, and impaired cognitive and learning-memory abilities. We observed that abnormalities in MRI/CT neuroimaging were consistent with brain hemorrhage and loss of brain parenchyma, which was confirmed by pathological analysis. Neuropathological findings included ventriculomegaly and cortical atrophy associated with impaired proliferation and migration of neural progenitor cells in the developing brain at both embryonic and postnatal stages. Robust inflammatory responses during infection were shown by elevated inflammatory cytokine levels, leukocyte infiltration, and activation of microglia and astrocytes in the brain. Pathological analyses and CT neuroimaging revealed brain calcifications induced by cMCMV infection and cell death via pyroptosis. Furthermore, antiviral treatment with ganciclovir significantly improved neurological functions and mitigated brain damage as shown by CT neuroimaging. These results demonstrate that this model is suitable for investigation of mechanisms of infection-induced brain damage and long-term studies of neurodevelopmental disorders, including the development of interventions to limit CNS damage associated with cCMV infection.

Many hearing difficulties can be explained as a loss of audibility, a problem easily detected and treated using standard audiological procedures. Yet, hearing can be much poorer (or more impaired) than audibility predicts because of deficits in the suprathreshold mechanisms that encode the rapidly changing, spectral, temporal, and binaural aspects of the sound. The ability to evaluate these mechanisms requires well-defined stimuli and strict adherence to rigorous psychometric principles. This project reports on the comparison between a laboratory-based and a mobile system’s results for psychoacoustic assessment in adult listeners with normal hearing. A description of both systems employed is provided. Psychoacoustic tests include frequency discrimination, amplitude modulation detection, binaural encoding, and temporal gap detection. Results reported by the mobile system were not significantly different from those collected with the laboratory-based system for most of the tests and were consistent with those reported in the literature. The mobile system has the potential to be a feasible option for the assessment of suprathreshold auditory encoding abilities.

Programmed cell death (PCD) plays a critical role in the development and maturation of the cochlea. Significant remodeling occurs among cells of the greater epithelial ridge (GER) of Kölliker’s organ, leading to tissue regression and formation of the inner sulcus. In mice, this event normally occurs between postnatal days 5–15 (P5-15) and is regulated by thyroid hormone (T3). During this developmental time period, the cochlea also contains a large population of macrophages. Macrophages are frequently involved in the phagocytic clearance of dead cells, both during development and after injury, but the role of macrophages in the developing cochlea is unknown. This study examined the link between developmental cell death in the GER and the recruitment of macrophages into this region. Cell death in the basal GER begins at P5 and enhanced numbers of macrophages were observed at P7. This pattern of macrophage recruitment was unchanged in mice that were genetically deficient for CX3CR1, the receptor for fractalkine (a known macrophage chemoattractant). We found that injection of T3 at P0 and P1 caused GER cell death to begin at P3, and this premature PCD was accompanied by earlier recruitment of macrophages. We further found that depletion of macrophages from the developing cochlea (using CX3CR1 DTR/+ mice and treatment with the CSF1R antagonist BLZ945) had no effect on the pattern of GER regression. Together, these findings suggest that macrophages are recruited into the GER region after initiation of developmental PCD, but that they are not essential for GER regression during cochlear remodeling.

The ability to perceive the world is not merely a passive process but depends on sensorimotor loops and interactions that guide and actively bias our sensory systems. Understanding which and how cognitive processes participate in this active sensing is still an open question. In this context, the auditory system presents itself as an attractive model for this purpose as it features an efferent control network that projects from the cortex to subcortical nuclei and even to the sensory epithelium itself. This efferent system can regulate the cochlear amplifier sensitivity through medial olivocochlear (MOC) neurons located in the brainstem. The ability to suppress irrelevant sounds during selective attention to visual stimuli is one of the functions that have been attributed to this system. MOC neurons are also directly activated by sounds through a brainstem reflex circuit, a response linked to the ability to suppress auditory stimuli during visual attention. Human studies have suggested that MOC neurons are also recruited by other cognitive functions, such as working memory and predictability. The aim of this research was to explore whether cognitive processes related to delayed responses in a visual discrimination task were associated with MOC function. In this behavioral condition, chinchillas held their responses for more than 2.5 s after visual stimulus offset, with and without auditory distractors, and the accuracy of these responses was correlated with the magnitude of the MOC reflex. We found that the animals’ performance decreased in presence of auditory distractors and that the results observed in MOC reflex could predict this performance. The individual MOC strength correlated with behavioral performance during delayed responses with auditory distractors, but not without them. These results in chinchillas, suggest that MOC neurons are also recruited by other cognitive functions, such as working memory.

Noise-induced hearing loss (NIHL) is one of the most prevalent forms of acquired hearing loss, and it is associated with aberrant microglial status and reduced hippocampal neurogenesis; however, the nature of these associations is far from being elucidated. Beyond its direct effects on the auditory system, exposure to intense noise has previously been shown to acutely activate the stress response, which has increasingly been linked to both microglial activity and adult hippocampal neurogenesis in recent years. Given the pervasiveness of noise pollution in modern society and the important implications of either microglial activity or hippocampal neurogenesis for cognitive and emotional function, this study was designed to investigate how microglial status and hippocampal neurogenesis change over time following acoustic exposure and to analyze the possible roles of the noise exposure-induced stress response and hearing loss in these changes. To accomplish this, adult male C57BL/6J mice were randomly assigned to either a control or noise exposure (NE) group. Auditory function was assessed by measuring ABR thresholds at 20 days post noise exposure. The time-course profile of serum corticosterone levels, microglial status, and hippocampal neurogenesis during the 28 days following noise exposure were quantified by ELISA or immunofluorescence staining. Our results illustrated a permanent moderate-to-severe degree of hearing loss, an early but transient increase in serum corticosterone levels, and time-dependent dynamic alterations in microglial activation status and hippocampal neurogenesis, which both present an early but transient change and a late but enduring change. These findings provide evidence that both the stress response and hearing loss contribute to the dynamic alterations of microglia and hippocampal neurogenesis following noise exposure; moreover, noise-induced permanent hearing loss rather than noise-induced transient stress is more likely to be responsible for perpetuating the neurodegenerative process associated with many neurological diseases.

Previous psychophysical studies have identified a hierarchy of time-averaged statistics which determine the identity of natural sound textures. However, it is unclear whether the neurons in the inferior colliculus (IC) are sensitive to each of these statistical features in the natural sound textures. We used 13 representative sound textures spanning the space of 3 statistics extracted from over 200 natural textures. The synthetic textures were generated by incorporating the statistical features in a step-by-step manner, in which a particular statistical feature was changed while the other statistical features remain unchanged. The extracellular activity in response to the synthetic texture stimuli was recorded in the IC of anesthetized rats. Analysis of the transient and sustained multiunit activity after each transition of statistical feature showed that the IC units were sensitive to the changes of all types of statistics, although to a varying extent. For example, we found that more neurons were sensitive to the changes in variance than that in the modulation correlations. Our results suggest that the sensitivity of the statistical features in the subcortical levels contributes to the identification and discrimination of natural sound textures.

Pathogenic variants in GJB2, the gene encoding connexin 26, are the most common cause of autosomal-recessive hereditary deafness. Despite this high prevalence, pathogenic mechanisms leading to GJB2-related deafness are not well understood, and cures are absent. Humans with GJB2-related deafness retain at least some auditory hair cells and neurons, and their deafness is usually stable. In contrast, mice with conditional loss of Gjb2 in supporting cells exhibit extensive loss of hair cells and neurons and rapidly progress to profound deafness, precluding the application of therapies that require intact cochlear cells. In an attempt to design a less severe Gjb2 animal model, we generated mice with inducible Sox10iCre ERT2 -mediated loss of Gjb2. Tamoxifen injection led to reduced connexin 26 expression and impaired function, but cochlear hair cells and neurons survived for 2 months, allowing phenotypic rescue attempts within this time. AAV-mediated gene transfer of GJB2 in mature mutant ears did not demonstrate threshold improvement and in some animals exacerbated hearing loss and resulted in hair cell loss. We conclude that Sox10iCre ERT2 ;Gjb2 flox/flox mice are valuable for studying the biology of connexin 26 in the cochlea. In particular, these mice may be useful for evaluating gene therapy vectors and development of therapies for GJB2-related deafness.

Failure in the structural maintenance of the hair cell stereocilia bundle and ribbon synapse causes hearing loss. Here, we have studied how ER stress elicits hair cell pathology, using mouse models with inactivation of Manf (mesencephalic astrocyte-derived neurotrophic factor), encoding an ER-homeostasis-promoting protein. From hearing onset, Manf deficiency caused disarray of the outer hair cell stereocilia bundle and reduced cochlear sound amplification capability throughout the tonotopic axis. In high-frequency outer hair cells, the pathology ended in molecular changes in the stereocilia taper region and in strong stereocilia fusion. In high-frequency inner hair cells, Manf deficiency degraded ribbon synapses. The altered phenotype strongly depended on the mouse genetic background. Altogether, the failure in the ER homeostasis maintenance induced early-onset stereociliopathy and synaptopathy and accelerated the effect of genetic causes driving age-related hearing loss. Correspondingly, MANF mutation in a human patient induced severe sensorineural hearing loss from a young age onward. Thus, we present MANF as a novel protein and ER stress as a mechanism that regulate auditory hair cell maintenance in both mice and humans.

Binaural hearing is critically important for the perception of sound spatial locations. The primary auditory cortex (AI) has been demonstrated to be necessary for sound localization. However, after hearing onset, how the processing of binaural cues by AI neurons develops, and how the binaural processing of AI neurons is affected by reversible unilateral conductive hearing loss (RUCHL), are not fully elucidated. Here, we determined the binaural processing of AI neurons in four groups of rats: postnatal day (P) 14–18 rats, P19–30 rats, P57–70 adult rats, and RUCHL rats (P57–70) with RUCHL during P14–30. We recorded the responses of AI neurons to both monaural and binaural stimuli with variations in interaural level differences (ILDs) and average binaural levels. We found that the monaural response types, the binaural interaction types, and the distributions of the best ILDs of AI neurons in P14–18 rats are already adult-like. However, after hearing onset, there exist developmental refinements in the binaural processing of AI neurons, which are exhibited by the increase in the degree of binaural interaction, and the increase in the sensitivity and selectivity to ILDs. RUCHL during early hearing development affects monaural response types, decreases the degree of binaural interactions, and decreases both the selectivity and sensitivity to ILDs of AI neurons in adulthood. These new evidences help us to understand the refinements and plasticity in the binaural processing of AI neurons during hearing development, and might enhance our understanding in the neuronal mechanism of developmental changes in auditory spatial perception.

Calcitonin-gene-related peptide (CGRP) is a lateral olivocochlear (LOC) efferent neurotransmitter. Depression of sound-driven auditory brainstem response amplitude in CGRP-null mice suggests the potential for endogenous CGRP release to upregulate spontaneous and/or sound-driven auditory nerve (AN) activity. We chronically infused CGRP into the guinea pig cochlea and evaluated changes in AN activity as well as outer hair cell (OHC) function. The amplitude of both round window noise (a measure of ensemble spontaneous activity) and the synchronous whole-nerve response to sound (compound action potential, CAP) were enhanced. Lack of change in both onset adaptation and steady state amplitude of sound-evoked distortion product otoacoustic emission (DPOAE) responses indicated CGRP had no effect on OHCs, suggesting the origin of the observed changes was neural. Combined with results from the CGRP-null mice, these results appear to confirm that endogenous CGRP enhances auditory nerve activity when released by the LOC neurons. However, infusion of the CGRP receptor antagonist CGRP (8–37) did not reliably influence spontaneous or sound-driven AN activity, or OHC function, results that contrast with the decreased ABR amplitude measured in CGRP-null mice.